The present invention is directed to novel compounds which act as prodrugs of AICA riboside and certain analogs of it. AICA riboside is a naturally occurring intermediate in purine biosynthesis. It is now known to enhance adenosine release from cells during ATP depletion. By virtue of its adenosine releasing abilities, AICA riboside has many therapeutic uses. However, AICA riboside does not cross the blood-brain barrier well and is inefficiently absorbed from the gastrointestinal tract; both characteristics decrease its full potential for use as a therapeutic agent.
Adenosine, 9-.beta.-D-ribofuranosyladenine (the nucleoside of the purine adenine), belongs to the class of biochemicals termed purine nucleosides and is a key biochemical cell regulatory molecule, as described by Fox and Kelly in the Annual Reviews of Biochemistry, Vol. 47, p. 635, 1978.
Adenosine interacts with a wide variety of cell types and is responsible for a myriad of biological effects. Adenosine serves a major role in brain as an inhibitory neuromodulator (see Snyder, S. H., Ann. Rev. Neural Sci. 8:103-124 1985, Marangos, et al., NeuroSci and Biobehav. Rev. 9:421-430 (1985), Dunwiddie, Int. Rev. Neurobiol., 27:63-130 (1985)). This action is mediated by ectocellular receptors (Londos et al., Regulatory Functions of Adenosine. pp. 17-32 (Berne et al., ed.) (1983)). Among the documented actions of adenosine on nervous tissue are the inhibition of neural firing (Phillis et al., Europ. J. Pharmacol., 30:125-129 (1975)) and of calcium dependent neurotransmitter release (Dunwiddie, 1985). Behaviorally, adenosine and its metabolically stable analogs have profound anticonvulsant and sedative effects (Dunwiddie et al., J. Pharmacol. and Exptl. Therapeut., 220:70-76 (1982); Radulovacki et al., J. Pharmacol. Exotl. Thera., 228:268-274 (1981)) that are effectively reversed by specific adenosine receptor antagonists. In fact, adenosine has been proposed to serve as a natural anticonvulsant, and agents that alter its extracellular levels are modulators of seizure activity (Dragunow et al., Epilepsia 26:480-487 (1985); Lee et al., Brain Res., 21:1650-164 (1984)). In addition, adenosine is a potent vasodilator, an inhibitor of immune cell function, an inhibitor of granulocyte oxygen free radical production, an anti-arrhythmic, and an inhibitory neuromodulator. Considering its broad spectrum of biological activity, considerable effort has been aimed at establishing practical therapeutic uses for adenosine and its analogs.
Sinc adenosine is thought to act at the level of the cell plasma membrane by binding to receptors anchored in the membrane, past work has included attempts to increase extracellular levels of adenosine by administering it into the blood stream. Unfortunately, because adenosine is toxic at concentrations that have to be administered to a patient to maintain an efficacious extracellular therapeutic level, the administration of adenosine alone is of limited therapeutic use. Further, adenosine receptors are subject to negative feedback control following exposure to adenosine, including down-regulation of the receptors.
Other ways of achieving the effect of a high local extracellular level of adenosine exist and have also been studied. They include: a) interference with the uptake of adenosine with reagents that specifically block adenosine transport, as described by Paterson et al., in the Annals of the New York Academy of Sciences, Vol. 255, p. 402 (1975); b) prevention of the degradation of adenosine, as described by Carson and Seegmiller in The Journal of Clinical Investioation, Vol. 57, p. 274 (1976); and c) the use of analogs of adenosine constructed to bind to adenosine cell plasma membrane receptors.
There are a large repertoire of chemicals that can inhibit the cellular uptake of adenosine. Some do so specifically, and are essentially competitive inhibitors of adenosine uptake, and others inhibit nonspecifically. P-nitrobenzylthioinosine appears to be a competitive inhibitor, while dipyridamole and a variety of other chemicals, including colchicine, phenethyalcohol and papaverine inhibit uptake nonspecifically.
Extracellular levels of adenosine can be increased by the use of chemicals that inhibit enzymatic degradation of adenosine. Previous work has focused on identifying inhibitors of adenosine deaminase, which participates in the conversion of adenosine to inosine. Adenosine deaminase activity is inhibited by coformycin, 2'-deoxycoformycin, and erythro-9-(2-hydroxy-3-nonyl) adenine hydrochloride.
A number of adenosine receptor agonists and antagonists have been generated having structural modifications in the purine ring, alterations in substituent groups attached to the purine ring, and modifications or alterations in the site of attachment of the carbohydrate moiety. Halogenated adenosine derivatives appear to have been promising as agonists or antagonists and, as described by Wolff et al. in the Journal of Biological Chemistry, Vol. 252, p. 681, 1977, exert biological effects in experimental systems similar to those caused by adenosine. Derivatives with N-6 or 5'-substitutions have also shown promise.
Although all three techniques discussed above may have advantages over the use of adenosine alone, they have been found to have several disadvantages. The major disadvantages of these techniques are that they rely on chemicals that have adverse side effects, primarily due to the fact that they must be administered in doses that are toxic, and that they affect most cell types nonselectively. As described in Purine Metabolism in Man, (eds. De Baryn, Simmonds and Muller), Plenum Press, New York, 1984, most cells in the body carry receptors for adenosine. Consequently the use of techniques that increase adenosine levels 9enerally throughout the body can cause unwanted, dramatic changes in normal cellular physiology. In addition, adenosine deaminase inhibitors prevent the degradation of deoxyadenosine which is a potent immunotoxin. [(Gruber et al. Ann. New York Acad. Sci. 451:315-318 (1985)].
It will be appreciated that compounds which increase extracellular levels of adenosine or adenosine analogs at specific times during a pathologic event, without complex side effects, and which would permit increased adenosine levels to be selectively targeted to cells that would benefit most from them, would be of considerable therapeutic use. By way of example, such compounds would be especially useful in the prevention of, or response during, an ischemic event, such as heart attack or stroke, or other event involving an undesired restricted or decreased blood flow, such as atherosclerosis or skin flap surgery, for adenosine is a vasodilator and prevents the production of superoxide radicals by granulocytes. Such compounds would also be useful in the prophylactic or affirmative treatmen of pathologic states involving increased cellular excitation, such as (1) seizures or epilepsy, (2) arrhythmias (3) inflammation due to, for example, arthritis, autoimmune disease, Adult Respiratory Distress Syndrome (ARDS), and granulocyte activation by complement from blood contact with artificial membranes as occurs during dialysis or with heart-lung machines. It would further be useful in the treatment of patients who might have chronic low adenosine such as those suffering from autism, cerebral palsy, insomnia and other neuropsychiatric symptoms, including schizophrenia. The compounds useful in the invention may be used to accomplish these ends.
Compounds which selectively increase extracellular adenosine would also be useful in the prophylactic protection of cells in the hippocampus implicated in memory. The hippocampus has more adenosine and glutamate receptors than any other area of the brain. Accordingly, as described below, it is most sensitive to stroke or any condition of low blood flow to the brain. Some recent studies suggest that Alzheimer's disease may result from chronic subclinical cerebral ischemia. The compounds of the invention, therefore, will be used for the treatment and/or prevention of both overt stroke and Alzheimer's disease.
It is now established that relatively short periods of brain ischemia (on the order of 2 to 8 minutes) set into motion a series of events that lead to an eventual death of selected neuronal populations in brain. This process is called delayed excitotoxicity and it is caused by the ischemia-induced release of the excitatory neurotransmitter glutamate. Within several days post-stroke the neurons in brain are overstimulated by glutamate to the point of metabolic exhaustion and death. Because glutamate appears to be the major factor involved in post-stroke cell damage, the blockade of glutamate receptors in brain could be beneficial in stroke therapy. In animals, glutamate receptor blockers have been shown to be effective in alleviating or reversing stroke associated neural damage. These receptor blockers have, however, been shown to lack specificity and produce many undesirable side effects. Church, et al., "Excitatory Amino Acid Transmission," pp. 115-118 (Alan R. Liss, Inc. 1987).
Adenosine has been shown to be a potent inhibitor of glutamate release in brain. The CA-1 region of brain is selectively sensitive to post-stroke destruction. In studies, where observations were made at one, three and six days post-stroke the CA-1 area was shown to be progressively destroyed over time. However, where cyclohexyladenosine ("CHA"), a global adenosine agonist, was given shortly after the stroke, the CA-1 area was markedly protected. (Marangos et al., Brain Res., in press.) That beneficial effect was also seen in the survival rate of the animals. Because of its global effect, however, CHA has non-specific side effects. For example it undesirably will lower blood pressure and could remove blood from the ischemic area, thereby causing decreased blood flow.
The compounds of the invention described and claimed herein not only show the beneficial adenosine release/glutamate inhibiting properties but are both site and event specific, avoiding the unwanted global action of known adenosine agonists.
Hyperglycemia has been reported to be associated with a poor prognosis for stroke. (Helgason, Stroke 19(8): 1049-1053 (1988). In addition, mild hypoglycemia induced by insulin treatment has been shown to improve survival and morbidity from experimentally induced infarct. (LeMay et al., Stroke 19(11): 1411-1419 (1988)). D-Ribose has been reported to cause hypoglycemia after oral or intravenous administration to experimental animals and humans and Foley (J. Clin. Invest. 37: 719-735 (1958) demonstrated an inhibition of phosphoglucomutase by ribose-5'-phosphate (formed intracellularly after ribose therapy). Although others have suggested that ribose lowers glucose via increased insulin release (Ishiwita et al., Endoncinol. Japan 25: 163-169 (1978)), the preponderance of evidence favors decreased glucose production ove increased utilization. AICA riboside and the prodrugs of the present invention could protect against ischemic injury to the central nervous system (CNS) by their ability to lower blood glucose.
Another area of medical importance is the treatment of neurological diseases or conditions arising from elevated levels of homocysteine (e.g., vitamin B12 deficiencies). The novel AICA riboside prodrugs of this invention may be used for such purposes as well.
A further area of medical importance is the treatment of allergic diseases, which can be accomplished by either preventing mast cell activation, or by interfering with the mediators of allergic responses which are secreted by mast cells. Mast cell activation ca be down-regulated by immunotherapy (allergy shots) or by mast cell stabilizers such as cromalyn sodium, corticosteroids and aminophylline. There are also therapeutic agents which interfere with the products of mast cells such as anti-histamines and adrenergic agents. The mechanism of action of mast cell stabilization is not clearly understood. In the case of aminophylline it is possible that it acts as an adenosine receptor antagonist. However, agents such as cromalyn sodium and the corticosteroids are not as well understood.
It will be appreciated, therefore, that effective allergy treatment with compounds which will not show any of the side effects of the above noted compounds, such as drowsiness in the case of the anti-histamines, agitation in the case of adrenergic agents, and Cushing disease symptoms in the case of the corticosteroids would be of great significance and utility. In contrast to compounds useful in the present invention, the AICA riboside prodrugs, none of the three known mast cell stabilizers are known or believed to be metabolized in the cell to purine nucleoside triphosphates or purine nucleoside monophosphates.
Clearly, there is a need for more effective anticonvulsant therapeutic compounds and strategies since most of the currently used antiseizure agents are toxic (e.g., dilantin), or are without efficacy in many patients. Adenosine releasing agents, which enhance adenosine levels during net ATP catabolism will be useful for the treatment of seizure disorders.
A concern in developing adenosine releasing agents, specifically AICA riboside, as anticonvulsants, however, resides in their less than full gastrointestinal tract penetration and their relatively low blood brain-barrier penetration. Derivatization of adenosine releasing agents, including AICA riboside has been undertaken with the goals of increasing penetration of AICA riboside into the brain and through the gut by delivering it as a brain and/or gut permeable form that avoids first pass metabolism and, while reaching the target regenerates into the parent compound (a prodrug strategy).
The present invention is directed to purine prodrugs and analogs which exhibit and, in some cases improve upon, the positive biological effects of AICA riboside and other adenosine releasing compounds without the negative effects of adenosine. The compounds herein defined may be used as prodrugs. The novel compounds typically exhibit one or more of the following improvements over AICA riboside: 1) more potent adenosine releasing effects; 2) increased half-lives; 3) increased brain penetration; 4) increased oral bioavailability; 5) increased myocardial targeting; 6) in some cases synergism with AICA riboside itself.
The AICA riboside prodrugs of this invention may be used in treatment and prevention of a number of disorders, some of which already have been mentioned.